Bottom Line:
In addition, we showed that lonafarnib treatment led to a dose-dependent decrease in scratch wound closure in vitro, whereas it had little effect on endothelial cell proliferation.These data indicate that lonafarnib inhibits neovascularization via directly targeting endothelial cells and disturbing their motility.Our findings thus offer novel mechanistic insight into the protective effect of farnesyl transferase inhibitors on atherosclerosis and provide encouraging evidence for the potential use of this group of agents in inhibiting plaque neovascularization.

ABSTRACTAtherosclerosis is a common cardiovascular disease that involves the build-up of plaque on the inner walls of the arteries. Intraplaque neovacularization has been shown to be essential in the pathogenesis of atherosclerosis. Previous studies showed that small-molecule compounds targeting farnesyl transferase have the ability to prevent atherosclerosis in apolipoprotein E-deficient mice, but the underlying mechanism remains to be elucidated. In this study, we found that lonafarnib, a specific inhibitor of farnesyl transferase, elicits inhibitory effect on vascular endothelial capillary assembly in vitro in a dose-dependent manner. In addition, we showed that lonafarnib treatment led to a dose-dependent decrease in scratch wound closure in vitro, whereas it had little effect on endothelial cell proliferation. These data indicate that lonafarnib inhibits neovascularization via directly targeting endothelial cells and disturbing their motility. Moreover, we demonstrated that pharmacological inhibition of farnesyl transferase by lonafarnib significantly impaired centrosome reorientation toward the leading edge of endothelial cells. Mechanistically, we found that the catalytic β subunit of farnesyl transferase associated with a cytoskeletal protein important for the establishment and maintenance of cell polarity. Additionally, we showed that lonafarnib remarkably inhibited the expression of the cytoskeletal protein and interrupted its interaction with farnesyl transferase. Our findings thus offer novel mechanistic insight into the protective effect of farnesyl transferase inhibitors on atherosclerosis and provide encouraging evidence for the potential use of this group of agents in inhibiting plaque neovascularization.

pone.0122830.g006: Lonafarnib suppresses MAPRE1 expression and interrupts its interaction with FTβ.(A) After co-transfection of GST-MAPRE1 and HA-FTβ, HUVECs were treated with DMSO or 10 μM lonafarnib for 24 hours. GST pull-down (PD) and Western blot were then performed to examine their interaction. The expressions of the HA-FTβwere monitored and comparable amounts of cell lysates were loaded in the pull-down assay (middle panel). (B) HUVECs were treated with DMSO or 10 μM lonafarnib for 24 hours, and endogenous MAPRE1expession was analyzed by Western blot.

Mentions:
Next, we sought to investigate whether lonafarnib affects the interaction of FTβwith MAPRE1. HUVECs were co-transfected with GST-MAPRE1 and HA-FTβ and then treated with DMSO or 10 μM lonafarnib for 24 hours. By GST pull-down assay, we found that lonafarnib significantly decreased the association between GST-MAPRE1 and HA-FTβ, compared with the control group (Fig 6A). Moreover, we also evaluated the effect of lonafarnib on the endogenous expression of MAPRE1. As shown in Fig 6B, we found that 10 μM lonafarnib treatment for 24 hours remarkably inhibited the expression of MAPRE1. Thus, these data showed that lonafarnib potently suppressed the expression of MAPRE1 and its association with FTβ.

pone.0122830.g006: Lonafarnib suppresses MAPRE1 expression and interrupts its interaction with FTβ.(A) After co-transfection of GST-MAPRE1 and HA-FTβ, HUVECs were treated with DMSO or 10 μM lonafarnib for 24 hours. GST pull-down (PD) and Western blot were then performed to examine their interaction. The expressions of the HA-FTβwere monitored and comparable amounts of cell lysates were loaded in the pull-down assay (middle panel). (B) HUVECs were treated with DMSO or 10 μM lonafarnib for 24 hours, and endogenous MAPRE1expession was analyzed by Western blot.

Mentions:
Next, we sought to investigate whether lonafarnib affects the interaction of FTβwith MAPRE1. HUVECs were co-transfected with GST-MAPRE1 and HA-FTβ and then treated with DMSO or 10 μM lonafarnib for 24 hours. By GST pull-down assay, we found that lonafarnib significantly decreased the association between GST-MAPRE1 and HA-FTβ, compared with the control group (Fig 6A). Moreover, we also evaluated the effect of lonafarnib on the endogenous expression of MAPRE1. As shown in Fig 6B, we found that 10 μM lonafarnib treatment for 24 hours remarkably inhibited the expression of MAPRE1. Thus, these data showed that lonafarnib potently suppressed the expression of MAPRE1 and its association with FTβ.

Bottom Line:
In addition, we showed that lonafarnib treatment led to a dose-dependent decrease in scratch wound closure in vitro, whereas it had little effect on endothelial cell proliferation.These data indicate that lonafarnib inhibits neovascularization via directly targeting endothelial cells and disturbing their motility.Our findings thus offer novel mechanistic insight into the protective effect of farnesyl transferase inhibitors on atherosclerosis and provide encouraging evidence for the potential use of this group of agents in inhibiting plaque neovascularization.

ABSTRACTAtherosclerosis is a common cardiovascular disease that involves the build-up of plaque on the inner walls of the arteries. Intraplaque neovacularization has been shown to be essential in the pathogenesis of atherosclerosis. Previous studies showed that small-molecule compounds targeting farnesyl transferase have the ability to prevent atherosclerosis in apolipoprotein E-deficient mice, but the underlying mechanism remains to be elucidated. In this study, we found that lonafarnib, a specific inhibitor of farnesyl transferase, elicits inhibitory effect on vascular endothelial capillary assembly in vitro in a dose-dependent manner. In addition, we showed that lonafarnib treatment led to a dose-dependent decrease in scratch wound closure in vitro, whereas it had little effect on endothelial cell proliferation. These data indicate that lonafarnib inhibits neovascularization via directly targeting endothelial cells and disturbing their motility. Moreover, we demonstrated that pharmacological inhibition of farnesyl transferase by lonafarnib significantly impaired centrosome reorientation toward the leading edge of endothelial cells. Mechanistically, we found that the catalytic β subunit of farnesyl transferase associated with a cytoskeletal protein important for the establishment and maintenance of cell polarity. Additionally, we showed that lonafarnib remarkably inhibited the expression of the cytoskeletal protein and interrupted its interaction with farnesyl transferase. Our findings thus offer novel mechanistic insight into the protective effect of farnesyl transferase inhibitors on atherosclerosis and provide encouraging evidence for the potential use of this group of agents in inhibiting plaque neovascularization.